Application of Process Raman Spectroscopy for bioprocessing using gene edited production hosts and cell-free systems

拉曼光谱(RS)是一种非破坏性的振动光谱技术，它提供了与水溶液中样品的化学或分子结构相关的尖锐光谱特征。生物分子、细胞和组织的拉曼测量已经被很好地建立，许多生物医学应用集中在疾病检测和体内血糖监测上。同时，RS在监测发酵过程方面也有了很大的发展。在现场，同时测量营养物质、代谢产物或副产物、细胞密度(或生物量)，使遥感成为工业生物处理中的重要过程分析技术。包括龙沙、葛兰素史克、默克-诺华-霍夫曼-拉罗氏在内的多家全球生物制造企业已经实施了在线拉曼光谱技术，用于过程控制和生物制造的优化。虽然基因组编辑在基因和细胞治疗应用中开辟了机会，但它也可以用于生物处理，以改善细胞生长特性；细胞生产率和产品滴度并优化和/或提高产品质量；改善作为生产宿主的无细胞提取物。细胞培养用于生物处理的另一种选择是使用基于细胞裂解物的无细胞蛋白表达(CPE)系统。最近，CPE已被证明是一种潜在的生物制品、疫苗和抗菌剂的生产宿主。CPE系统用于生物处理的优点是它们是非生物的，不容易受到污染，具有半限定的成分和一种在调整反应组合和表达过程方面提供灵活性的检测格式。拟议项目的总体目标是与设计和制造拉曼探针的工业合作者Kaiser(Endres-Hauser group)一起探索RS在新领域的应用，如基因组编辑、细胞培养和无细胞系统。拟议的项目汇集了弗里蒙集团在CPE和细胞培养方面的专业知识，哈萨克斯坦集团在RS方面的专业知识，以及伦敦生物铸造公司在合成生物学工具开发方面的专业知识。

Raman spectroscopy (RS) is a non-destructive vibrational spectroscopy technique that provides sharp spectral features that correlate to a sample's chemical or molecular structure in aqueous solution. Raman measurements of biological molecules, cells and tissues have been well established with many biomedical applications focused on disease detection and in vivo glucose monitoring. In parallel there has been substantial developments in the use of RS to monitor fermentation processes. In situ, simultaneous measurement of nutrients, metabolites or by-products, cell density (or biomass), have made RS an important Process Analytical Technology in industrial bioprocessing. A number of global biomanufacturers including for example Lonza, GSK, Merck Novartis Hoffman La Roche have already implemented online Raman Spectroscopy for process control and optimisation of biomanufacturing. Whilst genome editing has opened up opportunities in gene and cell therapy applications, it can also be used in bioprocessing to improve cell growth characteristics; cell productivity and product titre and optimise and/or improve product quality; improve cell free extracts as production hosts. An alternative to cell culture for bioprocessing is the use of cell-free protein expression (CPE) systems based on cell lysates. Recently CPE has been demonstrated as a potential production host for a variety of biologics, vaccines and antimicrobials. The advantages of CPE systems for bioprocessing it that they are non-living and not prone to contamination, have semi-defined components and an assay format that offer flexibility in adjusting the reaction mix and expression process. The overall aim of the proposed project is to explore the application of RS into new areas such as genome edited cell cultures and cell-free systems with an industrial collaborator Kaiser (Endres-Hauser group) who design and manufacture Raman probes. The proposed project brings together the expertise of Freemont group in CPE and cell culture, the Kazarian group in RS and London Biofoundry in synthetic biology tool development.